Cytokines play an important role in HIV pathogenesis. We have previously shown that treatment of monocytes with interferon-g (IFN-g), a cytokines produced by activated T lymphocytes, prior to exposure with HIV results in complete inhibition of HIV infection of monocytes. We have extended this study to obtain information on the mechanism(s) underlying IFN-g-induced inhibition of HIV infection of monocytes. We examined the effect of IFN-g on HIV entry into monocytes, the first event in the infectious cycle of the virus. Virus entry into monocytes was examined by PCR amplification of HIV-gag sequences in the genomic DNA and RNA of monocytes exposed to HIV after IFN-g treatment. The results from these experiments showed no evidence of either proviral DNA or HIV RNA sequences, and were consistent with the absence of intracellular virus particles either in latent or actively replicating state as determined by flow cytometric analysis of HIV-exposed monocytes. In addition, no HIV-induced cytopathic effects, such as multinucleated giant cell formation or cell death were observed in IFN-g-treated monocytes. Stimulation of IFN-g-treated monocytes 6 days post-infection with tumor necrosis factor-a (TNF-a), which is known to augment HIV replication in HIV-infected cells, did not result in the induction of HIV indicating the absence of latent HIV infection in IFN-g-treated monocytes. Treatment of monocytes with IFN-g, TNF-a, or with a combination of the two agents which is known to induce antimicrobial free radical nitric oxide (NO2-) in the murine system, did not induce NO2- production in human monocytes suggesting the antiviral activity of IFN-g to be independent of NO2--mediated killing of HIV or HIV-infected monocytes. IFN-g substantially down-regulated surface CD4 expression on monocytes and was directly related to intracellular HIV-p24 levels. These results indicate that IFN-g-mediated inhibition of HIV infection of monocytes was not due to virus latency or killing of these cells, but was primarily due to down-regulation of CD4 expression resulting in the inhibition of HIV entry into monocytes, and thus provide a mechanism of IFN-g-mediated inhibition of HIV infection of monocytes. Antiviral activity of IFN-g was also associated with the reduction of soluble factors that are involved in HIV pathogenesis and the inhibition of monocyte invasion through basement membrane matrices. Two manuscripts describing these studies are "in press". Further studies on the production of cytokines and other cellular factors are in progress to understand viral gene expression that may more clearly define the interplay of these events in the regulation of virus-cell interactions.